生理浓度睾酮对前列腺素F2α诱导SD大鼠血管平滑肌细胞中Ca^2＋升高的影响

Testosterone at Physiological Level Inhibits PGF_(2α)-induced Increase in Intracellular Ca^(2+) in Cultured Vascular Smooth Muscle Cells

目的:观察生理浓度睾酮短时作用对前列腺素F2α(PGF2α)诱导血管平滑肌细胞(VSMCs)中Ca2+升高的影响。方法:贴块法培养雄性SD大鼠胸主动脉VSMCs,钙敏荧光指示剂Fura-2负载细胞,Nikon TE-2000E活细胞工作站对VSMCs内钙信号进行测定。结果:VSMCs给以单纯PGF2α(10μmol/L)刺激,[Ca2+]i在数秒钟内由基线水平100nmol/L左右迅速升至约500nmol/L的峰值,随后呈缓慢下降,约150s降至基线水平。在PGF2α作用的峰值给以生理浓度(40nmol/L)睾酮或乙醇溶媒干预,前者能够明显缩短[Ca2+]i由峰值回落至基线水平的时间[与乙醇溶媒相比,(104.9±27.0)svs(153.5±40.4)s,P<0.01]。在PGF2α作用前预先给以睾酮或乙醇溶媒处理2min,前者对PGF2α刺激的[Ca2+]i峰值水平无明显影响[与乙醇溶媒相比,(390.0±126.0)nmol/Lvs(403.4±160.7)nmol/L,P>0.05],但使[Ca2+]i由峰值降至基线水平的时间显著缩短[与乙醇溶媒相比,(120.6±32.0)svs(151.4±27.4)s,P<0.01]。结论:睾酮对PGF2α诱导的VSMCs中[Ca2+]i升高具有快速抑制作用,提示睾酮通过与VSMCs细胞膜作用抑制受体门控钙通道介导的Ca2+内流。

Objective ： To explore the acute effects of testosterone at the physiological level on PGF2α-induced increase in intracellular Ca^2＋ in cultured vascular smooth muscle cells （VSMCs）. Methods ： VSMCs from the thoracic aorta of male Sprague-Dawley rats were cultured using the explant method. The subconfluent VSMCs were incubated with serum-free medium for 24 hours to obtain quiescent non-dividing cells and then treated with the indicated agents. For the measurement of [ Ca^2＋ ] i, the VSMCs were loaded with fura-2. Changes of [ Ca^2＋ ] i were determined ratiometrically with a Nikon TE-2000E system. Results ： The resting level of [ Ca^2＋ ] i was around 100 nmoL/L in the VSMCs. Exposing cells to perfusate containing 10 μmol/L PGF2α triggered an immediate and transient peak in [ Ca^2＋ ] i , which gradually decreased afterwards. Interference at the peak with the physiological concentration （40 nmol/L） of testosterone significantly decreased the peak-to-baseline time of [ Ca^2＋ ]i, compared with ethanol vehicle （104.9 ±27.0 s vs 153.5±40.4 s, P 〈 0.01 ）. Pretreatment with testosterone at 40 nmol/L for 2 minutes also reduced the peak-to-baseline time of [ Ca^2＋ ] i significantly in comparison with the ethanol control （ 120.6 ± 32.0 s vs 151.4± 27.4 s, P 〈 0. 01 ）, but it had no significant effect on the peak level of PGF2α-induced intracellular Ca^2＋ （390.0 ± 126.0 nmol/L vs 403.4 ±160.7 nmol/L, P 〉 0.05）. Conclusion： Testosterone at physiological concentration inhibits PGF2α-induced Ca^2＋ fluxes, probably via receptor-operated calcium channels by a non-genomic mechanism in VSMCs, which may be involved in the vasodilatory effect of testosterone.